Fragment 31-35 of beta-amyloid peptide induces neurodegeneration in rat cerebellar granule cells via bax gene expression and caspase-3 activation - A crucial role for the redox state of methionine-35 residue

The amyloid beta-peptide (A beta P) is the major protein component of brain senile plaques in Alzheimer's disease. The redox state of methionine-35 residue plays a critical role in peptide neurotoxic actions. We used the fragment 31-35 of A beta P [A beta P(31-35)], containing a single methionine-35 residue (Met-35), to investigate the relationship between the oxidative state of Met-35 and neurotoxic and pro-apoptotic actions induced by the peptide; in rat cerebellar granule cells (CGC), we compared the effects of A beta P(31-35), in which the Met-35 is present in the reduced state, with those of a modified peptide with oxidized Met-35 [A beta P(31-35)Met-35(OX)], as well as an A beta P-derivative with Met-35 substituted by norleucine [A beta P(31-35)Nle-35]. A beta P(31-35) induced a time-dependent decrease in cell viability. A beta P(31-35)Met-35(OX) was significantly less potent, but still induced a significant decrease in cell viability compared to control. No toxic effects were observed after treatment with A beta P(31-35)Nle-35. A beta P(31-35) induced a 2-fold increase in bax mRNA levels after 4 h, whereas A beta P(31-35)Met-35(OX) raised bax mRNA levels by 41% and A beta P(31-35)Nle-35 had no effect. Finally, A beta P(31-35) caused a 43% increase in caspase-3 activity after 24 h; A beta P(31-35)Met-35(OX) caused only a 18% increase, and A beta P(31-35)Nle-35 had no effect. These findings suggest that A beta P(31-35)-induced neurodegeneration in CGC is mediated by a selective early increase in bax mRNA levels followed by delayed caspase-3 activation; the redox state of the single Met-35 residue is crucial in the occurrence and extent of the above phenomena.